1. Field of the Invention
This invention relates generally to antisense oligonucleotides and specifically to the enhancement of cell death by treatment of the cell with an antisense oligonucleotide specific for an anti-apoptotic gene in combination with a therapeutic means for induction of cell death.
2. Background of the Prior Art
Apoptosis is a form of cell death in which the cell actively participates in its own demise. Apoptosis is defined predominantly by morphological criteria. The characteristic features of apoptosis include cell shrinkage, chromatin condensation, and DNA fragmentation into oligonucleosomal ladder size units. Finally, fragments of the dying cell form sealed vesicles called apoptotic bodies which are rapidly removed by neighboring cells (Wylie, et al., Int. Rev. Cytol., 68:251, 1980).
Most approaches to cancer therapy result in activation of apoptotic pathways. For example, apoptosis is the major form of cell death associated with the action of chemotherapeutic agents on tumor cells. Expression of genes that interfere with apoptosis can have important consequences for the efficacy of therapeutic approaches. In certain circumstances, drug resistance in cancer cells is actually due to resistance to apoptosis in general. Therefore, an antiapoptotic signal might provide such resistance to therapeutic approaches aimed at inducing cell death. For example, the expression of the bcl-2 gene in tumor cells has been shown to increase the resistance of cells to cytotoxic agents (Lozzio, et al., Blood, 45:321, 1975). It would be highly desirable to develop a method to identify those genes that interfere with apoptosis in order to regulate the expression of those genes such that the cells would become more susceptible to the induction of apoptosis and, therefore, enhance the efficacy of therapeutic treatment of the cells.
Resistance to chemotherapeutic treatment has been observed in chronic myelogenous leukemia (CML) cells. CML is characterized cytologically by the Philadelphia (Ph) chromosome, which results from the translocation of the c-abl gene on chromosome 9 to the bcr gene on chromosome 22. The result of this translocation is the expression of a bcr-abl fusion protein. Despite the fact that the function of the bcr-abl fusion protein has been studied by different groups, drug resistance of the CML patient to chemotherapeutic treatment remains an unsolved problem. Szczylik, et al., (Science, 253:562, 1991) recently indicated that antisense oligonucleotides to the bcr-abl breakpoint junction inhibited cell proliferation in patient-derived CML cells, and suggested using such antisense alone to treat CML patients. More recently, Szczylik, et al., (J. Clin. Invest. 92:194, 1993) also showed that treatment of Philadelphia leukemic cells, mixed 1:1 with normal bone marrow cells, with a combination of a low dose of mafosfamide and antisense to the bcr-abl breakpoint junction, was effective in killing the leukemic cells while sparing a high number of the normal cells and at least part of the abl sequence. The authors also emphasized the importance of not including antisense to the bcr region in order to avoid perceived deleterious side effects. However, based on this therapeutic approach, therapeutic antisense oligonucleotides would have to be individually designed in order to be effective, since the breakpoint junction between bcr and abl appears to vary from patient to patient.
Therefore, there is a need to develop a generalized method for inducing apoptosis in a cell having a bcr-abl translocation which avoids the necessity of determining a patient's particular bcr-abl breakpoint nucleotide sequence in order to develop an effective therapeutic agent. The present invention provides such a method and therapeutic agents.